MGN-3 Research Data

MGN-3

Journal Articles: 

EFFECT OF MGN-3 ON HUMAN NATURAL KILLER CELL ACTIVITY AND INTERFERON- y SYNTHESIS IN VITRO.

1 - MGN-3-

MGN-3 is an arabinoxylane from rice bran that has been enzymatically modified by Hyphomycetes mycelia. It is polysaccharides that contain ß -1.4 xylopyronase hemicellulose. MGN-3 was offered by Daiwa Pharmaceutical Co.. Ltd. Tokyo. Japan.

2 - Separation of peripheral blood Lymphocytes (PBL):

Mononuclear cells from the subjects were separated from the fresh whole blood by ficollhypague density gradient centrifugation (Litton Bionetics, Rockville. MD). The lymphocyte band at the interface was collected and cells pelleted by centrifugation, washed twice in RPM11640 and suspended in complete medium (CM) which consists of RPMI-1640, supplemented with 10% (v/v) fetal calf serum. 2mM glutamine, 25 mM Hepes (pH 7.2). 50 units penicillin and 50 ug streptomycin per ml (Grand Island Biologicals. Santa Clara. CA).

3 - In Vitro Treatment of PBL with MGN-3:

PBL were cultured with MGN-3 at different concentration (0.25, 50 and 100 ug/ml for 16 hr. PBL was washed twice with CM and adjusted at I OA- 106 cells I ml.

4-NK Cell Activity:

The most common method of assessing NK anti-Cancer activity is to use Cr-release assay in which fixed number of Chromium, (CR)- labeled tumor target cells are incubated with varying number of NK effector cells. The percent cell mediated lympholysis is calculated from the foillowing equation:

% lysis = Exp. Rel. - Sp- Rel X 100 Total Rel. - Sp. Rel

Where Exp. Rel test refers to the counts in supernatants from wells containing effector cells. Sp. Rel is the spontaneous release in medium only, and Total Rel. is the maximum counts obtained by lysis with detergent. Usually four serial dilution are made of the NK effector cells and plated in quadruplicate with the tumor targets in microtiter plates. In brief. 5 x 101 per ml CM. and 0.1 pipetted to quadruplicate wells to give and effector: Target cell ratio of 100: 1, 50:1, 25:1 and 1 2:1 . Microtiter plates were then centrifuged for 3 minutes at 500 RPM and following a 4 hours incubation at 37C. and the supernatant were counted in a gamma counter. The percentage of isotope released was calculated by the formula mentioned above.

5 - MTT Asaay

Effect of MGN-3 on P388 tumor cell proliferation was examined by MTT assay. 0.1 n-d -of tumor cells 2xlOl cultured with MGN-3 different doses in each well of 96 well microtiter plate rounded bottom for one day. 10 li I of MTT (3-(4.5-dimethylthazol-2-yl)- 2.5 diphenyl tetrazolium bromide) were added to each well for four hours and the plate centrifuged for 10 min. at 2000 RPM. Supernatant was discarded. and 100 ml of acid alcohol was added to each well. The microtiter plate was read by MR 700 machine.

6 - Assay of I FN- -Y Ploduction:

Peripheral blood lymphocytes (I X 106 CelIS/Ml) were incubated with various concentrations of MGN-3 (0-200 Mg/ml) for 48 Hr. at 37C under 50% CO, and supernatants were collected for assay of IFN- y production. The amount of IFN- y in the supernatants was determined by ELISA using a commercially available IFN- y ELISA kit from BioSource International (Camarillo. CA).

Results of this study demonstrated that.

1. Treatment with MGN-3 resulted in significant increase in NK activity post treatment (Figure 1)

2. The mechanism by which MGN-3 augments NK activity through induction of IFN- y (Figure 2).

3. MGN-3 also possesses direct anti-cancer activity in vitro. (Figure 3). We Conclude that:

1. MGN-3 is a potent Biological Response Modifier (BRM) as manifested by highly increase in human NK cell activity at 16 hr. post exposure.

2. Earlier work in vivo demonstrated significant increase in human NK cell activity by MGN-3 at 2 wks post treatment.

3. The mechanism by which MGN-3 induce NK activity is through production of IFN-'Y 

4. MGN-3 possesses direct anti-cancer activity in vitro.

5. The induction of NK activity by MGN-3 may represent a new immune-therapeutic approach for the treatment of cancer. which needs to be examined in multiple clinical trials.
 
 

EFFECT OF MON-3 ON HUMAN NATURAL KILLER CELL ACTIVITY 
AND INTERFERON-y SYNTHESIS IN VITRO.

M.Ghoneum. G. Namatalla and C.Kim. Drew University of Medicine and Science, Los Angeles, CA 90059, and University of California, fivinc,CA.92717

MGN-3 is an extract of arabinoxylane from rice bran that has been enzymatically treated with an extract from hyphomycetes mycelia. In this study we investigated the effect of MGN-3 on natural killer (NK) cell activity and interferon- -y (IFN-,y ) synthesis by peripheral blood mononuclear cells (MNC). MNC was prepared from peripheral blood of healthy individuals and incubated with various concentrations of MGN-3 for 16 hrs, and then NK cell activity was measured by 4 hr "Cr-release assay using K562 Tumor cells as target NK activity was significantly enhanced (2-5 fold) by treatment with MGN-3 at 0-100 p glml. In an attempt to investigate the mechanism by which MGN-3 enhances NK activity, we examined the effect of MGN-3 on IFN- y production by MNC. Culture supernatants of the cells incubated with MGN-3 were collected and analyzed for INF- -y synthesis by ELISA. UIF- y production was increased>2 fold. We concluded that MGN- 3 is a potent biological response modifier (BRM) and may be useful in immunotherapy of cancer.

MGN-3. was offered by Daiwa Pharmaceutical Co., Ltd. Tokyo-Japan.

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